Reverse genetic approaches are a powerful laboratory tool for determining the function of a target protein. The target protein is “knocked down,” and cellular changes are observed in order to infer the normal function of the knocked down target. Methods for knocking down a target protein by manipulating DNA transcription and RNA translation are well established. Among the most commonly used are gene knockout in whole animals and degradation of target mRNA using siRNA and shRNA techniques. However, delivery of RNA molecules can be cumbersome, and these methods often cannot achieve 100% efficiency.
Although several methods exist for knocking down a protein target at the transcription and translation levels, there are very few options for knocking down a protein target once the protein has been made. Such methods are desirable because most small molecule therapeutics operate by manipulating proteins directly. Therefore, targeted degradation techniques that operate at the protein level provide the best tool for examining the potential effects of small molecule therapeutics.
The ideal protein knock down system would function at the protein level, and would be capable of tightly controlling protein levels in a temporal manner. Temporal control of protein levels allows down-regulation of proteins that are essential for the full development of a system or pathway, and can be used to study dynamic biological functions which are otherwise difficult to manipulate.